Abstract The microflora of the anterior male urethra during adolescence is poorly described and no data address the range of typical urethral microflora during adolescence as functions of pubertal development or onset of various types of partnered sexual activity. This means that current understanding of the urethral microbiome in adolescent men rests entirely on data drawn from adults, and from epidemiologic and behavioral explanations. Marked ethnic group variation in adolescent sexual behaviors and disparity in conditions such as sexually transmitted infections (STI) justifies exploration of potential ethnic group differences in the male urethral microflora. Some research among adult males suggests important interactions between the male urethral microflora and STI, but this research has not been extended to younger men. Here we propose to characterize the male urethral microbiome in longitudinal urine samples collected from a multi-ethnic cohort of adolescent males. Microbial flora will be assessed as function of pubertal development, sexual activity, and incident STI. Subjects will be Latino (N=24), African-American (N=24), and Euro-American (N=24) males, ages 14 - 17 at enrollment; total N=72). Participants will be recruited from an urban clinic within an area of high prevalence of STI. Socio-demographic data will be collected at enrollment. Daily behavioral and symptomologic data will be collected via cell phone, which will also be used for cohort maintenance and scheduling of sample collections. Urine will be collected at enrollment and up to 36 1-month follow-up samples for characterization of microflora using high-throughput 16s rRNA phylogeny, Q-PCR and metagenomic sequencing. STI testing will be done as well to better understand interactions between the urethral microbiome and infection. Cell phones also will be used to initiate daily sample collection for 10 days following trigger events such as specific sexual acts, antibiotic use, condom failure, or genital symptoms.